Methods for screening for transdominant effector peptides and RNA molecules

Abstract

Methods and compositons for screening for transdominant effector peptides and RNA molecules selected inside living cells from randomized pools are provided.

We claim:  
     
       1. A method of screening for a transdominant bioactive agent that alters the phenotype of a cell, said method comprising the steps: 
       a) introducing a molecular library of randomized candidate nucleic acids, each operably linked to nucleic acid encoding a secretion signal, into a first plurality of cells, wherein each of said nucleic acids comprises a different nucleotide sequence, wherein said randomized candidate nucleic acids are expressed in said first cells to produce a plurality of randomized peptides; and  
       b) screening a second plurality of cells which are co-cultured with said first plurality of cells for a cell exhibiting an altered phenotype, wherein said altered phenotype is due to the presence of a transdominant bioactive agent.  
     
     
       2. A method according to  claim 1  further comprising the step: 
       c) isolating said cell exhibiting an altered phenotype.  
     
     
       3. A method according to  claim 2  further comprising the step: 
       d) isolating a candidate nucleic acid from said cell exhibiting an altered phenotype.  
     
     
       4. A method according to  claim 2  or  3  further comprising the step: 
       e) isolating a target molecule using  
       i) a candidate nucleic acid; or  
       ii) the expression product of a candidate nucleic acid.  
     
     
       5. A method according to  claim 1  wherein said candidate nucleic acids are each operably linked to a nucleic acid encoding a fusion partner. 
     
     
       6. A method according to  claim 5  wherein said fusion partner comprises a presentation sequence that presents said expression product in a conformationally restricted form. 
     
     
       7. A method according to  claim 1  wherein said introducing is with retroviral vectors. 
     
     
       8. A method according to  claim 1  wherein at least one plurality of said first and second plurality of cells are mammalian cells. 
     
     
       9. A method according to  claim 1  wherein said library comprises at least 10 4  different nucleic acids. 
     
     
       10. A method according to  claim 1  wherein said library comprises at least 10 5  different nucleic acids. 
     
     
       11. A method according to  claim 1  wherein said library comprises at least 10 6  different nucleic acids. 
     
     
       12. A method according to  claim 1  wherein said library comprises at least 10 7  different nucleic acids. 
     
     
       13. A method according to  claim 1  wherein said library comprises at least 10 8  different nucleic acids.